Fig. 4

Lamellar protrusions at the tip of the ES exist in open and closed configurations.

(A) Select images from serial-section scanning electron microscopy of a 5.5 dpf zebrafish’s right inner ear. Dorsal is up, lateral left, medial right, ventral down, anterior top, and posterior bottom of the series z-stack. Cells forming lamellar barriers were labeled (color overlays, consistent across panels) to highlight connectivity of lamellae. Presented slices are a subset from the series, each separated by 960 nm (Video 8). The lumen of the ES is labeled and outlined with a white dashed line in third panel; perilymph (peri.). (B) Lamellae interdigitate and can form tongue-in-groove structures (inset). (C) Lamellae can interweave. (D) Example of lamellar junctions in an open configuration. (E) Cells in endolymphatic duct have basal lamellae, with the presented duct connecting with ES in panel A. (F) 3D rendering of ES segmentation from serial micrographs shown in panel (A) and Video 8. Black dotted-outline encompasses area of closed, endolymph-filled lamellae. Black mesh highlights areas of membrane overlap between lamellae that are spread open. (G) Electron-dense tight junctions (yellow arrows) present in cells that also have spread basal protrusions. An opening in the apical junctions creates a path from the duct to the basal protrusions (magenta arrow, slices in three panels 1.2 μm apart). (H, H’) lmx1bb^jj410/jj410 embryos maintain apical junctions between ES cells (yellow arrows). (I) Mutant ES cells lack basal protrusions (red arrow). Scale bar in (A) is 1000 nm, (F) is 5 μm, and all other scale bars are 500 nm. (B–G) Serial section electron microscopy, n = 2, additional transmission EM, n = 3. (H–I) Serial section EM of mutant, n = 1, transmission EM, n = 3.